Sialidases ganglioside substrates

In addition to these methods, new techniques based on the principle of enzyme-linked immunosorbent assay (ELISA) have recently been introduced for sialidase assay. A glycoprotein or ganglioside substrate is coated in plastic microtiter wells and incubated with an enzyme preparation, followed by detection of the desialylated product using an appropriate ligand. Thus, the activity of Clos-... 

Sialidase activities directed toward ganglioside substrates in plasma membrane and lysosomal fractions can be distinguished based on their responses to detergents and inhibitors. The requirement of detergents significantly differs between both enzyme activities (see Section 5.1). Heparin or heparin sulfate almost completely inhibits the GM3 sialidase activity in the plasma membrane preparation, but only slightly affects the lysosomal enzyme activity. The inhibition of the plasma membrane enzyme, but not the lysosomal enzyme, can be... 

Ohman and Svennerholm (1971) measured the appearance of sialidase activity toward ganglioside substrate in developing human brain. Activity was first detected in the fetal brain at 15 to 20 weeks and by term had increased to half of the adult level. During the first year of life sialidase activity may decrease slightly, and then it increases until it approaches the adult level at 5 years. The appearance of sialidase activity in the later stages of development may reflect the need for the enzyme to alter, or to help in degradation of, the more complex sialyl compounds synthesized, and these latter in turn may serve to induce sialidase activity. 

The substrate-specificity studies described in this Subsection show striking similarities between the sialidases from various sources, but also characteristic differences with regard to the N- and O-substitu-tions of Neu, the sialic acid glvcosidic linkages, and the oligosaccharide, glycoprotein, or ganglioside nature of the substrates.55... 

Variable sialidase activities are known to occur physiologically during ontogenesis."0 In chick retina, the lowest enzyme activity was observed in the 8-day-old embryo, whereas the highest activity was measured at hatching. Gangliosides were shown to be the best substrates... 

Exposure of HeLa cells to butyrate had no effect on the activity of GM3-sialidase when GM3 specifically labeled in the sialic acid residue was used as substrate (Fig. 3a). We were unable to detect any "ecto"-sialidase activity in either control or butyrate-treated cells (14) although others have postulated that such an enzyme is important in regulating plasma membrane gangliosides (15,16). In contrast, the activity of the specific sialyl transferase involved in GM3 biosynthesis increased over 20-fold following butyrate treatment (Fig. 3b). The effect was specific as activities of the other glycosphingolipid transferases that could be measured in HeLa cells were not altered in butyrate-treated cells (4,8,17). 

C. Oehler, J. Kopitz, and M. Cantz, Substrate specificity and inhibitor studies of a membrane-bound ganglioside sialidase isolated from human brain tissue, J. Biol. Chem., 383 (2002) 1735-1742. 

---